The proposed SCOR will test two basic hypothesis to explain the CF-related modifications in airway surface liquid (ASL) that lead to colonization by Pseudomonas. The serous cell malfunction hypothesis proposes that serous cells in the submucosal glands of large airways and on the surface of small airways show reduced secretion of fluid and antibiotics in CF. The resulting decreased levels of serous cell antibiotics and increased mucin concentrations in ASL favor pathogen colonization. The high salt hypothesis proposes that human airway epithelium absorbs Na and Cl, that a substantial fraction of Cl absorption occurs by an transcellular route, and that block of this route by malfunction of CFTR promotes higher than normal NaCl content of ASL in CF. A less likely possibility is that elevated NaCl levels in CF ASL are caused by saltier than normal gland secretions resulting from failure of ductal salt absorption. High salt content of ASL may encourage Pseudomonas colonization by inhibiting the action of natural antibiotics. Project 1 (Widdicombe/Bastacky) will use low-temperature scanning electron microscopy and X-ray microanalysis of rapidly frozen tissues to determine how the regulation of ASL depth is altered in CF. Salt and mucin content of ASL and gland changes in depth, composition and viscosity of ASL in living tissues using novel fluorescence microscopy. Project 3 (Wine) will study the secretory mechanisms contributing to the ASL, and test specifically the serous cell malfunction hypothesis. Project 4 (Miller) will determine the routes and mechanisms by which Na and Cl are absorbed across airway surface epithelium. Nasal PD measurements will be used to verify in vitro findings. A Cell Culture Core (Finkbeiner) will provide intact human airways and cultures of human gland and surface epithelial cells.